Chiral stationary phase based on yohimbine

ABSTRACT

A set of chiral stationary phases is based on yohimbine and its derivatives. In particular, the hydroxyl functionality of yohimbine and its analogs is covalently bonded via a urethane linkage to a polymethylenesilyl chain attached to the bound hydroxyl groups of a refractory inorganic oxide by Si-O bonds. The resulting chiral stationary phases have multiple chiral recognition sites and can be used with a broad spectrum of materials as eluents without leaching.

BACKGROUND OF THE INVENTION

Ever since Pasteur discovered the property of optical activity displayedby chiral compounds, the resolution of racemic mixtures into theirenantiomeric components has posed a challenge. Substantial progress inseparating enantiomeric pairs has been achieved since Pasteur'slaborious hand separation of the enantiomeric crystals of racemic sodiumammonium tartrate, yet methods of resolution, and the materials usedtherefor, remain a formidable obstacle to commercial production ofoptically active organic substances.

A traditional method of resolution comprises reacting a racemic mixturewith a second optically active substance to form a pair ofdiastereomeric derivatives. Such derivatives generally have differentphysical properties which permit their separation by conventional means.For example, fractional crystallization often permits substantialseparation to afford at least one of the diastereomers in a pure state,or largely so. An appropriate chemical transformation then converts thepurified derivative, which was formed initially solely to prepare adiastereomeric pair, into one enantiomer of the originally racemiccompound. This traditional method is exemplified by the reaction ofnaturally occurring optically active alkaloids, for example, brucine,with racemic acids to form diastereomeric salts, with release of anoptically active organic acid from a purified diastereomer uponacidification of the latter.

Such traditional methods suffer from many limitations. Generally, onlyone of the enantiomeric pairs can be obtained, so yields are necessarilyless than 50%. The separation of the material so obtained usually isincomplete, leading to materials with enhanced rather than completeoptical purity. The optically active materials used to form thediastereomers frequently are expensive and quite toxic--the alkaloids asa class are good examples-and are only partially recoverable.Regeneration of optically active material from its derivative may itselfcause racemization of the desired compound, leading to diminution ofoptical purity. For example if optically active benzyl alcohols areprepared through their diastereomeric ester derivatives, subsequent acidhydrolysis of the latter to regenerate the alcohol may be accompanied byappreciable racemization.

With the advent of chromatography diverse variations on the basic themeof separating diastereomers became possible. These approaches undeniablyrepresent substantial advances in the art, yet fail to surmount thebasic need, and associated problems, to prepare diastereomericderivatives of the desired compound and to transform such derivativesafter separation to the optically active compounds of interest.

Chromatographic methods of separating diastereomers offer advantages ofgeneral application, mild conditions which generally preclude chemicalor physical transformation, efficiency of recovery and separation whichare limited only by the number of theoretical plates employed and thecapability of utilization from a milligram to kilogram scale.Translation from a laboratory to industrial scale has proved feasible,and commercial processes employing chromatographic separation occupy animportant position in the arsenal of available industrial methods. Forsuch reasons, methods based on chromatographic separation remain underintensive exploration.

To circumvent the disadvantage of separating diastereomeric derivativesof a compound while retaining the advantage of chromatographicseparation, recent advances in the art have employed chiral, opticallyactive compounds in association with the chromatographic support. Thetheory underlying this approach is that chiral material will havedifferential weak interactions with enantiomers, for example, hydrogenbonding, or acid-base interactions generally. Such weak interactionslead to reversible formation of entities which we refer to as complexes,and the equilibrium constant characterizing complex formation will bedifferent for each member of the enantiomeric pair. The differentequilibrium constants manifest themselves as a differing partitioncoefficient among the phases in a chromatographic process, leadingultimately to separation of enantiomers.

Thus, enantiomers of some chromium complexes were resolved bychromatography on powdered quartz, a naturally occurring chiralmaterial. Karagounis and Coumolos, Nature, 142, 162 (1938). Lactose,another naturally occurring chiral material, was used to separatep-phenylene-bis-iminocamphor. Henderson and Rule, Nature, 141, 917(1938). However, despite this knowledge substantiating theoreticalconsiderations, advances in the art have been tortuous at best.

A major obstacle has been development of a chiral solid phase capable ofresolving, at least in principle, a broad class of racemic organiccompounds, with a stability which permits repeated usage, and withadequate capacity to make separation feasible on a preparative scale.Gil-Av has made a major contribution toward one kind of solution bygas-liquid phase chromatographic resolution of enantiomers using columnscoated with N-trifluoroacetyl derivatives of amino acids, di-andtri-peptides. Gil-Av and Nurok, "Advances in Chromatography", Volume 10,Marcel Dekker (New York), 1974. However, the advances suffer practicallimitations originating from the need to have volatile substrates andthe inability to scale up the methods employed.

Another advance is represented by the work of Baczuk and coworkers, J.Chromatogr., 60, 351 (1971), who covalently bonded an optically activeamino acid through a cyanuric acid linkage to a modified dextran supportand utilized the resulting material in column chromatography to resolve3,4-dihydroxyphenylalanine. A different approach is exemplified bypolymerization of optically active amides with the resulting polymerused as a solid phase in liquid-solid chromatography. Blaschke andSchwanghart, Chemische Berichte, 109,1967 (1976).

More recently it has become an accepted reality that enantiomericmedicinals may have radically different pharmacological activity. Forexample, the (R)-isomer of propranolol is a contraceptive whereas the(S)-isomer is a beta-blocker. An even more dramatic and tragicdifference is furnished by thalidomide where the (R)-enantimer is a safeand effective sedative when prescribed for the control of morningsickness during pregnancy whereas the (S)-enantiomer was discovered tobe a potent teratogen leaving in its wake a multitude of infantsdeformed at birth. This has, in part, provided the motivation fordeveloping additional tools for chiral separations. Chromatographicprocesses, especially liquid chromatography, appear to offer the bestprospects for chiral separations. One variant of the latter utilizesachiral eluents in combination with chiral stationary phases (CSPs),which has the critical aspect that a variety of chiral stationary phasesbe available to the practitioner. In recent years substantial progresshas been made by developing a class of chiral stationary phases basedupon derivatized polysaccharides, especially cellulose, adsorbed on acarrier such as silica gel or a modified silica gel. This recently hasbeen summarized by Y. Okamoto, J. Chromatog., 666 (1994), 403-19.

However effective may be the aforedescribed supports based onpolysaccharides, there remains a need for chiral stationary phases wherechirality is imparted by a monomer rather than by oligomers or polymersas represented by the polysaccharides. To be optimally useful the chiralmonomer should have a plurality of chiral sites, so as to offer severalchiral recognition sites and afford the potential of being broadly usedin chiral separations. An appropriate monomer should afford a CSP basedon covalent linkage of the monomer to the underlying carrier; covalentlyattaching the chiral monomer to a carrier virtually eliminates leaching,regardless of the mobile phase, and permits one to use many more typesof mobile phases and to switch from forward to reverse phase eluentsusing the same column without fear of destroying the CSP due to leachingor plugging of the column. This benefit makes such CSPs much moreeffective for traditional single pass chromatography, for recycle-typechromatography, for simulated moving bed-based chromatography, andsimple preferential adsorption of one enantiomer over the other.

The use of a monomeric chiral host containing several chiral centersproviding a plurality of potential chiral interactions offers thepossibility of a chiral stationary phase manifesting broad chiraldiscrimination. Yohimbinic acid is a chiral material with several easilyderivatizable sites making this chiral host readily modifiable to "tune"its selectivity according to the racemate to be resolved. Covalentattachment of yohimbinic acid to the underlying carrier via itscarboxylic acid function affords a useful series of chiral stationaryphases, but use of the carboxylic acid functionality as the site ofattachment does have some unwanted features. Modeling indicates that thechiral sites are more hindered by the surface of the carrier whenyohimbinic acid is attached via the acid moiety as opposed to itsattachment via the hydroxyl moiety. While some steric hindrance isdesirable as a means to promote chiral selectivity, too much hindrancewill actually limit access of both enantiomers to the active chiralsites, leading to decreased chiral recognition and poorer separations.Such hindrance will also decrease the number of different types ofracemates which the chiral support can separate.

Attaching the yohimbinic structure to the carrier via the hydroxyl groupinstead of the acid moiety offers a slightly different chiral surface tothe racemates. This variation increases the versatility of theyohimbinic acid structure, and it may also increase the number ofpossible racemates which may be separable. However, there are possibledrawbacks to simply attaching yohimbinic acid to the carrier via thehydroxyl group. One is that the free acid moiety is likely to interferewith any reaction designed to couple the hydroxyl group to the carrier.Even if such a reaction is successfully carried out, there is a freecarboxylic acid moiety available to interact with any racemateapproaching the chiral discriminator since the carboxylic acid moiety isthe most polar group in yohimbinic acid. In other chiral stationaryphases where chiral recognition occurs using polar interaction, freecarboxylic acid sites may significantly hinder, if not completelynegate, chiral selectivity. Therefore, if the yohimbinic structure is tobe attached to a carrier, it is desirable and perhaps imperative thatthe carboxylic acid moiety be blocked, as by forming an alkyl ester.Yohimbine is the methyl ester of yohimbinic acid.

In this application we take advantage of covalent attachment ofyohimbinic acid esters, whose primary example is yohimbine, to theunderlying carrier via the free hydroxyl group. The use of this monomershould lead to chiral stationary phases with good mass transferproperties more similar to brush-type stationary phases, whereas CSPsbased on high carbon-loaded derivatized cellulosics show impaired masstransfer properties. Yohimbine-based CSPs according to our inventiondescribed within may be expected to be effective in both analytical andpreparative chromatography, especially simulated moving-bedchromatography.

SUMMARY OF THE INVENTION

The purpose of our invention is to prepare a variety of covalentlybonded chiral stationary phases based on yohimbine and analogous estersmanifesting broad chiral discrimination. An embodiment is yohimbine oran analogous ester of yohimbinic acid covalently bonded to an underlyingsilica carrier via an alkylsilyl spacer. A specific embodiment of thisvariant is one where a urethane group formed from the hydroxyl group ofyohimbine and an isocyanato group is the covalent link bonding yohimbineto the spacer molecule. In a more specific embodiment the CSP is theyohimbine urethane of propylsilanized silica. Other embodiments will beapparent from our ensuing description.

DESCRIPTION OF THE INVENTION

The need for broadly-effective, "general-purpose" chiral stationaryphases reflects the need for chiral stationary phases having 1) anorganic monomer as the chiral recognition agent and 2) the potential tohave broad chiral discrimination associated with a plurality of chiralsites. Because a CSP with covalently bound organic material can beexpected to be more stable toward leaching and to afford somewhatgreater flexibility in operating conditions, this type of CSP often ispreferred over one where the organic material is merely coated on acarrier Our invention fills these needs by using yohimbine and itsanalogs (i.e., the esters of yohimbinic acid) as the chiral organicmaterial with a multiplicity of chiral recognition centers. Yohimbineand its analogs are covalently bound to the underlying carrier via analkylsilyl spacer. Because yohimbine has multiple functionality, severalsites may be derivatized independently to alter and customize chiralrecognition for optimum resolution of specific enantiomeric pairs.

The chiral stationary phases of our invention consist of a carrier,which is a refractory inorganic oxide, and yohimbine or an analogthereof, where the yohimbine or analog thereof is covalently bound tothe carrier via a spacer.

The carriers of our invention are refractory inorganic oxides whichgenerally have a surface area of at least about 35 m² /g, preferablygreater than about 50 m² /g and more desirably greater than 100 m² /g.There appears to be some advantage to working with materials having ashigh a surface area as possible, although many exceptions are knownwhich preclude making this a general statement. Suitable refractoryinorganic oxides include alumina, titania, zirconia, chromia, silica,boria, silica-alumina and combinations thereof. Of these, silica isparticularly preferred as a carrier in chromatographic separations.Since the chiral stationary phase is yohimbine or an analog thereofcovalently bonded to the underlying carrier, it is required that thecarrier have bound surface hydroxyl groups, so that the latter may formone end of a tether which results from reaction of the bound surfacehydroxyl groups with a silane functionality on a compound to form acovalent OSi bond as part of the structure, carrier-OSi-spacer. Theprogenitor of the spacer portion of our invention has the formula(AO)_(x) SiHal_(y) (CH₂)_(n) --NCO. The silane part of our spacerprogenitor contains either halogen, Hal, or alkoxy groups, AO, eitheralone or in combination. Chlorine is by far the most common halogenwhich may be used in the practice of our invention, although brominealso may be used equally well. As for the alkyl group of AO, A may beany alkyl group, but preferably is a lower alkyl having from 1 throughabout 6 carbon atoms, with 1 and 2 carbon alkyl groups particularlydesirable. The silicon atom is separated from the nitrogen atom by achain of methylene groups, CH₂. The length of this chain is given by nwhich is an integer between 2 and about 10, with n=2 to 4 especiallydesirable. The subscripts x and y also are integers where their sum isequal to 3. A suitable progenitor of the spacer portion of our inventionmerely has a non-reactive group between the silicon atom and thenitrogen of the isocyanato group. Thus, another equally viableprogenitor of the spacer portion of our invention is one where themethylene chain is replaced by an aromatic group, as in ##STR1## Othersuitable progenitors will be apparent to the skilled worker.

Yohimbine and its analogs in all cases constitute the chiral organicmaterial in the chiral stationary phase of our invention. Forconvenience, yohimbine itself is given by the formula, (R₁ =CH₃, R₂ =H).##STR2## One notes that yohimbine contains two centers other than thehydroxyl group which are easily substituted or derivatized, giving riseto the variables R₁ and R₂. R₁ is selected from the group consisting ofalkyl moieties containing from 1 up to about 20 carbon atoms, and aryland aralkyl moieties containing from 7 up to about 20 carbon atoms,although the variant where R₁ is a lower alkyl group containing 1through about 6 carbon atoms is favored. R₂ is selected from the groupconsisting of hydrogen, alkyl moieties containing from 1 up to about 20carbon atoms, alkylaminocarbonyl moieties having 2 to 10 carbon atoms,arylaminocarbonyl moieties having 6 to about 10 carbon atoms, and acylmoieties containing from 2 up to about 20 carbon atoms.

Covalent bonding to the underlying carrier occurs via the hydroxylportion of the yohimbine. A generalized representation of the resultingcovalently bonded chiral stationary phase is given below: ##STR3## Thegroups R₁ and R₂ have been defined above. The yohimbine or analogthereof may be present in an amount from about 0.2 up to about 8 wt. %based on the finished chiral stationary phase. Formation of the CSPresults from the reaction of the isocyanate group in anisocyanotoalkylsilyl material with the hydroxyl functionality inyohimbine or an analog thereof to form a urethane linkage. Whether theurethane is first formed with later reaction of the silyl functionalitywith the bound surface hydroxyl groups of the carrier, or whether theurethane is formed after reaction of the silyl functionality with thebound surface hydroxyl groups of the carrier, is a matter of choice andis not fundamental to the outcome of our inventions. These two variantscan be represented by the following equations (where ZOH representsyohimbine and its analogs). ##STR4##

The example which follows merely illustrates some specific embodimentsof our invention, which is not limited thereto. Other variants andembodiments will be clear to the skilled artisan.

EXAMPLE 1

Chiral Stationary Phase Based on (+)-Yohimbine: To a 100 mL,three-necked, round-bottomed flask equipped with a reflux condenser, athermometer (attached to a Therm-o-watch temperature controller), aTeflon-coated stirring bar, and a heating mantle, were added 1.00 g(2.821 mmol) of (+)-yohimbine (the methylester of (+)-yohimbinic acid)and 40 mL of a mixture of dry pyridine and benzene. To the top of thecondenser were attached a 10 mL equilibrated dropping funnel and anitrogen line. Into the dropping funnel was added 0.734 g (2.821 mmol)of 3-isocyanatopropyltriethoxysilane (95%, Huls America) dissolved inabout 10 mL of dry pyridine. The flask contents were stirred, heated to80° C., and the isocyanate was slowly added over a 15 minute period. Thebenzene was distilled from the reaction until the temperature reached90° C., then the reaction was allowed to proceed for about 24 hoursmore. The reaction progress may be followed by FT-IR.

After 24 hours, the contents (now containing the urethane product fromthe reaction of the hydroxyl moiety of the yohimbine with the isocyanategroup of the organosilane) were stripped of a portion of the pyridine.The pyridine removed was replaced with dry benzene. Stripping may becarried out using a stream of dry nitrogen or by pouring the contentsinto a 100 mL, single-necked, round-bottomed flask and stripping thepyridine from the reaction mixture using a rotary evaporator (set at 85°C.) and reduced pressure. The residue was returned to the same 100 mLreaction apparatus, which was equipped as before except the droppingfunnel was removed and a Dean-Stark trap was added between the flask andthe condenser. The nitrogen line was attached to the top of thecondenser.

To the reaction residue were added 60 mL of benzene followed by 4.00 gof 5 μ silica gel. The slurry was gently stirred and the reactionmixture brought to reflux. Periodically, about 20 mL of benzene wereremoved from the trap and replaced with fresh, dry benzene. At the endof 16 hours, the reaction was stopped and the contents filtered on a 60mL (M) sintered glass funnel. The filter cake was washed sequentially(3×30 mL) with pyridine, acetone, methanol, acetone, and pentane thenair dried in the funnel. The modified silica gel was fully dried in avacuum oven at 5 torr for 3 hours at about 60° C. to yield the chiralstationary phase as a powder.

I claim as my invention:
 1. A chiral stationary phase represented by##STR5## where carrier represents a refractory inorganic oxide havingbound surface hydroxyl groups, O-Si is the covalent bond between thebound surface hydroxyl groups of said refractory inorganic oxide andsilicon, where R₁ is selected from the group consisting of alkylmoieties containing from 1 up to 10 about 20 carbon atoms, and aryl andalkaryl moieties containing from 7 up to about 20 carbon atoms, andwhere R₂ is selected from the group consisting of hydrogen, alkylmoieties containing from 1 up to about 20 carbon atoms,alkylaminocarbonyl moieties having two to 10 carbon atoms,arylaminocarbonyl moieties having 6 to about 10 carbon atoms, and acylmoieties containing from 2 up to about 20 carbon atoms.
 2. The chiralstationary phase of claim 1 where the refractory inorganic oxide issilica.
 3. The chiral stationary phase of claim 1 where .
 4. The chiralstationary phase of claim 1 where R₁ is methyl and R₂ is hydrogen. 5.The chiral stationary phase of claim 1 where R₁ is an alkyl having from1 up to about 6 carbon atoms.
 6. A chiral stationary phase comprising acarrier of a refractory inorganic oxide covalently bonded via boundsurface hydroxyl groups to silicon atoms contained in a spacer agent offormula (RO)_(x) Hal_(y) Si(CH₂)_(n) NHC(O)--, where R is an alkylgroup, Hal is a halogen, x and y are integers such that x+y=3, and n isan integer from 1 up to about 12, and where said spacer agent iscovalently bonded at the carbon atom of the carbonyl group to thehydroxyl group of yohimbine and its analogs.